1. Technological Field to which Invention Belongs
The present invention relates to a liquid pipetting apparatus, which dispenses and pipettes a liquid of small volume, for example, liquids such as reagents and molten metals, and an array manufacturing apparatus, which manufactures the array constituted by arranging the spot of the liquid including a probe on a substrate in a matrix shape with high density.
2. Relating Technology
Survey instruments of a hemanalysis machine, a genetic screening, and a pharmaceutical screening, etc. are requested to reduce the volume of the waste fluid for the running cost reduction. The volume of the waste fluid is chiefly constituted from two of reactive test solutions and washing water for reactive containers, so that the volume of a reactive test solution should be made reduced in order to reduce this waste fluid. A reactive test solution is decided according to a minimum dispense volume of a liquid dispensing device, so that it is effective to make a minimum dispense volume a minute volume for the reduction in volume of the waste fluid.
Moreover, these survey instruments are required to decrease the inspection time in order to make an increase in volume of the inspection and the inspection item in recent years correspondent. It is effective to decrease the required time to the chemical reaction process of a reactive test solution to shorten the inspection time, but to this end, a minimum dispense volume should be made a minute volume, and the constructed reactive test solution should be made minimized. In addition, micro array attracts attention as a new genetic screening technology in recent years. Micro array is a chip which utilized to arrange the liquid of a liquid sample containing the fragment of various micro array on the substrate in the matrix shape, to react it with the reagent and to observe the reactive result with the fluorometry or the like. For the micro array, a liquid sample can be arranged on the substrate with a high density by making the volume of a minimum charging of a liquid sample a minute volume, as a result, a many of samples can be analyzed at a time, so that the minute volume making of the volume of a minimum charging leads to shortening the analysis time.
In this way, it is required to make a minimum dispense volume a minute volume for reducing the volume of the waste fluid and shortening the inspection time in various survey instruments and analyzer used for the medical treatment field.
In the past, the syringe piston pump has been used as a liquid dispense technology of such a survey instrument. However, there is a problem that the volume of a minimum dispense of the syringe piston pump is about 2 μL, and the dispense precision becomes deteriorated extremely when the liquid of small volume or more from this was dispansed. As for such a problem, a nanopipette, which applies the ink jet method as a technology for reducing the volume of a minimum dispense by two digits or more with comparison to the conventional method, is shown in for example Japanese Patent Application Opened No. 8-233710.
FIG. 44 shows a schematic block diagram to which the main portion of the nanopipette in the ink jet method is shown by the cross-section.
In FIG. 44, this main portion comprises a pipette chamber 152 formed in a pipette frame 151, a nozzle 153 communicated to the atmosphere, for generating and dispensing the liquid, a reservoir 154, a piezo-electric element 156 to supply driving force to the pipette chamber 152 through a diaphragm 155, a controller 157 for controlling the piezo-electric element, and Ian introduction entrance 158 where reagent or sample is introduced into the reservoir 154.
In the conventional embodiment, an apparatus, which makes a reactive sample of the liquid by mixing a specimen sample and a necessary reagent, comprises a pipette element capable of generating the liquid by which the volume, which does not exceed 0.1 nl, is assumed to be a unit, there is proposed a proposal adopting an ink jet method which performs a pipetting of a reagent and a reactive sample necessary to construct a liquid reactive sample with an volume which does not exceed least 1 nl and with a unit which does not exceed resolution of 0.1 nl.
The dispensing speed of this pipette can be raised in proportion to the frequency of the high-frequency wave by which the acceleration power is driven, for example, in case of driving it by 10 kHz, the construction of one reaction sample can be achieved in case of driving it with 10 kHz in the maximum for example in about 1 sec. per 1 μl. As a result, a reactive sample can be constructed with high speed such as in 100 samples per one minute-two minutes.
However, in the prior art, the acceleration applying element such as piezo-electric elements gives liquid the acceleration power directly or through the partition such as diaphragms indirectly, so that in the case that the piezo-electric element touches the liquid directly, there is a problem that the feature of washing the conduit becomes difficult since a liquid sample intrude in the crystal grain field on the surface of the piezo-electric element. Moreover, in case of having the partition such as diaphragms, there is a problem that the structure comes to complicate, and as a result, the manufacturing cost rises.
Moreover, in the case that the bubble is generated in the chamber with reference to the evaporation of the cavitation and dissolved oxygen etc., the problem that the pressure generated by the acceleration applying element attenuates with the bubble, and the liquid cannot be charged, generates.
In addition, the feature of making plural arrays of pipettes can became possible by adopting the above system. That is, a chamber, a reservoir, and driving element are small size in the pipette of the system, and the voltage is only applied to the piezo-electric element whose drive mechanism is also a driving element, so that the controlling mechanism is also simple and can also drive a plural driving elements easily, and independently. By mounting the arrayed pipette to a suitable movement element, and driving the piezo-electric element as driving element of a necessary pipette portion while properly changing a relative position with a reactive container where plural reactive holes are arranged on the plane according to necessary timing, the feature of achieving the pipette of the object reagent became possible with high throughput for a reactive hole at the object position so as to just draw by the ink-jet printer.
The hemanalysis machine, the genetic screening, and the pharmaceutical screening or the like are measured by making a liquid sample which contains a chemical compound, blood and DNA or the like react chemically, and by measuring the result of the reaction with the use of the fluorometry and the spectrophotometer or the like. Therefore, there is a problem that when impurities influencing on the chemical reaction are mixed with the liquid sample, an accurate inspection result can not be obtained.
In order to prevent such a problem, there is devised that the dispensing device is used as disposable, and the dispensing device is changed in case of dispensing a different kind of liquid sample. If such a system is adopted, impurities might not mix with the conduit. However, the inspection machine of the pharmaceutical screening etc. handles the kind of liquid sample of several thousand from several hundreds, so that the method of exchanging the dispensing device whenever the liquid sample is changed, is not realistic in plane of the cost. Moreover, the prior art uses the piezo-electric element as an actuator, but if the influence on the environment is considered since the piezo-electric element contains a large volume of lead, the feature of using the dispense apparatus as disposable becomes a problem. In order to resolve these problems, the actuator and the conduit are constructed detachably, if only the conduit may be exchanged when the liquid sample is exchanged the component to be abandoned is only the conduit, so that the cost becomes cheap and the adverse effect is not caused for the environment.
Here, in the prior art, a method of separating and detaching the actuator and the conduit is considered. In the prior art, there is utilized that the actuator of the piezo-electric element etc. supplies the acceleration power to the liquid directly, or through the shroud element such as diaphragms indirectly. In the structure that the actuator gives liquid the acceleration power directly, the actuator and the conduit cannot naturally be separated detached. On the one hand, in the structure to use the partition element such as diaphragms, if some installation and deinstallation elements to separate both between the actuator and the diaphragm are provided, the feature of detaching the actuator and the conduit becomes possible. However, the diaphragm has very thin shape to make a small displacement of few μm act on the liquid, and has the problem of easily arriving at the deformation or the breakage by little power which causes in the case of detaching works. Even if the diaphragm does not damage, the position of the diaphragm is easily displaced before and behind the detachable working due to the size error between the assembly error which causes at the detachable works and the exchanged member, as a result, the problem causes that the acceleration power, which acts on the liquid, changes and the dispense volume changes.
Moreover, in a liquid dispensing device with the use of the ink jet system, the smaller the nozzle diameter is to generate a small liquid, the more advantageous, the nozzle diameter should be made φ 0.1 mm or less to generate the liquid of at most 0.1 nl or less as in the prior art. On the other hand, the genetic screening machine and the biochemistry analysis machine or the like often contain the cell and the protein in the handled liquid sample. These liquid samples come to generate the nozzle clogging easily according to the experience of the inventor when the nozzle diameter becomes φ0.1 mm or less.
Moreover, in the case where the nozzle blocking is generated, by performing following process that the nozzle is soaked in the washing water or the like and the pressure is raised by supplying the liquid is supplied in the chamber, the work for removing the blocked substance is performed, however, there is a case by which the substance blocked in the nozzle cannot be removed even if such restoration work is done, too. Moreover, in a case that when the substance blocked in the nozzle is removed, the nozzle is damaged, the problem that the dispense volume changes before and after the nozzle blocking, may be happen, too. In case where such a problem occurs, only the nozzle has to be able to be exchanged, but in the prior art, the nozzle, the chamber, and the actuator are constituted to one body, so that the nozzle alone cannot be exchanged, and the entire dispensing device should be exchanged, and thus this invited the problem as an increase in the running cost and an increase of waste, etc.
Moreover, the liquid is supplied from the introduction entrance to the reservoir in the prior art, but the case, that the liquid is attracted from the nozzle, is not described, however, when liquid in little volume of many kinds of samples are inspected, as in the hemanalysis machine and the genetic screening machine, etc., aspirating and the dispensing systems are preferable from the viewpoint of the inspection treatment power. In the prior art, in the case of attracting the sample from the nozzle, the aspiration method is considered by installing the liquid aspiration means such as pumps behind the introduction entrance. However, a long time is required to attract the liquid from the nozzle where the caliber is as small as the prior art, so that there is a problem that the time required for the inspection becomes long, too as a result.
On the other hand, if the conduit of the dispensing device can be cleanly washed, many kinds of liquid samples can be dispensed without exchanging the conduit by the same dispensing device. In the prior art, though a general washing is performed by pouring the washing water in the conduit, but it is difficult for the flow rate in the vicinity of the pipette chamber and the reservoir to flush the sample which adheres to the surface of the conduit extremely late since the caliber of the nozzle is small even if it tries to pour the washing water from the reservoir side in the conduit.
The prior art of the manufacturing method of the micro array (probe array) is disclosed in Japanese Patent Application Laid-open No. 187900/1999. In the prior art, the spot including the probe is formed on a solid phase by adhering the liquid sample which contains the probe as the liquid drop to the solid phase (solid substrate) by the ink jet method. Hereafter, the outline of the manufacturing method of the array according to the above prior art is explained by using FIG. 45 and FIG. 46 which is the cross-sectional view on the A—A line of FIG. 45.
In FIG. 45, numeral 161 is a liquid supply system (nozzle) which can dispensably hold the liquid including probe (for example nucleic acid probe) as the dispensing liquid, numeral 163 is solid phase (for example transparent glass board) to which said nucleic acid probe should be combined, numeral 165 is a kind of the ink jet head and is a bubble jet head including a mechanism, which gives the thermal energy to the liquid and dispenses the liquid, and numeral 164 is a liquid dispensed from the bubble jet head 165 and including the nucleic acid probe.
Moreover, in the bubble jet head 165 shown in FIG. 46, numeral 167 is a liquid including the nucleic acid probe to be dispensed, and numeral 177 is a substrate section having the heat portion for giving the dispense energy to the liquid. The substrate section 177 includes a protective coat 169 formed with oxidation silicon or the like, electrodes 171-1 and 171-2 formed with aluminum or the like, a heat resistive layer 173 formed with Nichrome wire or the like, a heat storage layer 175, and a supporting body 176 formed with alumina or the like having excellent heat radiative property.
The liquid 167 containing the above nucleic acid probe, reaches a dispense orifice 179 (dispense vent), and forms meniscus 181 with the given pressure. Under such a condition, when an electric signal is applied to electrodes 171-1 and 171-2, the region (foam region)shown by a sign 183 is heated rapidly and generates heat, so that the bubble is generated in the liquid 167 touched to this region. The meniscus is dispensed by the pressure of the bubble, and liquid 167 is dispensed from the orifice 179. The dispensed liquid 167 flies toward the surface of the solid phase 163.
The volume of the liquid capable of being dispensed with the bubble jet head including the above structure is different according to the size or the like of the nozzle, but this volume can be controlled in the order of, for example, 4–50 picoliter, so that it is extremely effective as the means to arrange nucleic acid probe with high density. Moreover, in the official gazette of the above prior art, it is described that it can use the piezo jet head for dispensing the liquid in the nozzle according to utilizing the vibrating pressure of the piezo element, in place of the above bubble jet head.
The above prior art has the advantage by which the probe sample of a minute volume can be arranged on the solid phase substrate with high accuracy and with high density, but the other bubbles may be mixed in the liquid in the dispense head in addition to the bubble for dispensing the liquid, and thus in the case that the bubble mixes in the dispense head, there is a case that the volume of the dispensing liquid is changed and the dispense of the liquid can not be performed. In that case, the problem is caused that the spot diameter of the array becomes an ununiformity. That is, in the above prior art, there is a case that the bubble mixes in the conduit, occasionally, by exchanging the liquid sample and by evaporating dissolved oxygen in the liquid sample, so that in case of mixing the bubble, pressure for dispensing the liquid becomes attenuated by the mixed bubble, and thus the volume of the dispensing liquid changes.